Alcohol abuse and dependence affect an estimated 8.5% of the U.S. population and are responsible for substantial health and societal costs. Multiple lines of evidence have shown that activation of the extrahypothalamic corticotropin-releasing factor (CRF) system plays a critical role in the negative emotional state and enhanced motivation to drink experienced by ethanol-dependent subjects. A key question that remains unanswered so far is the source of CRF being released in the central nucleus of the amygdala (CeA) during ethanol withdrawal. Filling this gap of knowledge is a prerequisite for investigating the transcriptional mechanisms inducing CRF upregulation in ethanol dependence, and possibly identifying novel targets for the treatment of alcoholism. In the present proposal, we will test th hypothesis that CeA CRF neurons directly contribute to the emotional disruption characterizing ethanol withdrawal and to the escalation of ethanol drinking in dependent mice. A subhypothesis is that at least some of those neurons are intrinsic to the CeA and may therefore be responsible for increased CRF release in the CeA during ethanol withdrawal. To test these hypotheses, we will use transgenic mice injected with viral vectors to selectively target the population of CeA neurons expressing CRF. The first Specific Aim will assess whether some of those neurons are intrinsic to the CeA, using both neurotracing and electrophysiological approaches. In the second Specific Aim, in vivo silencing of CRF expression and excitation of CRF neurons in the CeA will address the behavioral relevance of CeA CRF neurons to the symptomatology of ethanol withdrawal, using paradigms of anxiety-like behavior and voluntary drinking. The proposed experiments are expected to establish the functional implication of CeA CRF neurons in the emotional and motivational aspects of ethanol withdrawal. They will pave the way for future investigation of the molecular mechanisms driving the recruitment of the CRF system in alcoholism and other stress-related psychiatric disorders.